Most protocols used in biochemistry and molecular biology comprise multiple steps that must be executed properly and in the proper order. Exemplary steps include mixing, diluting, centrifuging, separating, extracting, suspending, heating, cooling, reacting, dispensing, and the like. Many of these steps are similar, at least in part, in various protocols. A principal difference of one protocol versus another is often the particular order in which the steps are executed. Another difference is a different value of a parameter (e.g., temperature or reaction time) in one protocol versus another. Yet another difference is the inherent variability of different people performing nominally similar steps.
An example of a molecular biological protocol involving a defined series of steps, in which most of the steps are repeated multiple times, is the Polymerase Chain Reaction (PCR) protocol. Since its inception, PCR has been modified and tailored for use in multiple specific situations. Consequently, although there are currently multiple PCR protocols, many of the steps in them are substantially the same because, for example, substantially all existing protocols involve performing multiple temperature “cycles.” This has generated interest in automating PCR protocols. In addition, there have been attempts to derive PCR protocols that use progressively smaller sample and reagent volumes and that can perform a PCR cycle in less time.
For example, there have been various attempts to incorporate the complete process of PCR assays into microchannels, using microfluidics, to reduce sample volumes. Since a microchannel is a fluid passageway formed with a specific configuration, it cannot be changed easily to accommodate departures from the particular protocol for which the microchannel was configured. Consequently, a key problem with this approach is that the user cannot change the assay protocol easily. Also, the small size of the microchannel device complicates isolating the different locations at which respective thermal steps in the PCR cycle are conducted, and prevent the removal and characterization of sample aliquots at any stage in the PCR cycle.
Decreasing the sample size inevitably involves the production and manipulation of small droplets (e.g., “microdroplets”) of liquid. In the various efforts at automating laboratory procedures, manipulating small droplets has been the focus of much attention in recent years. Berthier and Silberzan, Microfluidics for Biotechnology, Artech House, Norwood, 2006. Use of small droplets allows significantly smaller reaction volumes and decreased assay times. The two primary modes of conventional droplet manipulations are: (1) manipulating discrete liquid plugs in pre-defined microchannels (Belder, Angew. Chem. Int. Ed. 44:3521-3522, 2005; Joanicot and Ajdari, Science 309:887-888, 2005), and (2) manipulating liquid droplets resting on an open, flat surface (Mugele and Baret, J. Phys: Condens. Matter 17:R705-R774, 2005; Su et al., ACM Trans. Design Autom. Electron Syst. 11:442-464, 2006). The first mode (1) incorporates “liquid plug” techniques and the second mode (2) incorporates “open-surface” techniques. Changing an automated liquid-plug technique is difficult essentially because changing the technique involves changing the microfluidic plumbing. Open-surface techniques, involving the manipulation of liquid droplets on an open, flat surface, has been demonstrated most notably in association with magnetofluidics, in which the droplets containing paramagnetic particles move over a hydrophobic surface under the influence of an external magnetic field. Egatz-Gomez et al., Appl. Phys. Lett. 89:034106, 2006. A difficulty with this technique is that paramagnetic particles have not yet been designed that do not interfere with biological reactions. Another technique in this general category is called electrowetting on dielectrics (“EWOD”). Cho et al., J. Microelectromech. Syst. 12:70-80, 2003. Although this technique allows precise droplet movement, splitting, and merging, an apparatus performing this method is comparatively difficult to fabricate and operate, and has problems with diffusional mixing and contamination from increased wetting on the surface. Thus, a continuing need exists for improved systems and methods of manipulating biological liquids in a microfluidic environment.